Lung epithelial progenitor cells are essential for development and adult airway regeneration and repair. The mechanisms controlling their behaviors including proliferation and differentiation during development and the progression of airway injury repair are largely unknown. MicroRNAs (miRNAs) are crucial modulators of gene expression, yet their involvement as regulators of lung epithelial progenitor cells is still poorly understood. Recently, We have demonstrated that miR-302/367 is a direct target of Gata6 -a transcription factor essential for proper lung development and airway regeneration- and regulates the balance of lung epithelial progenitor cell proliferation and differentiation as well s cell apical-basal polarity. We have begun to explore the importance of miR-302/367-mediated lung epithelial progenitor development. Overexpression of miR-302/367 in developing airway epithelium causes expansion of Sox2+ proximal and Sox9+ distal progenitor cells and decreased airway epithelial cell differentiation. Notably, our recent studies reveal that expression of miR- 302/367 is markedly increased in naphthalene-injured lungs compared to control uninjured lungs, suggesting a potential role for miR-302/367 in mediating lung epithelial gene transcription and the injury response during airway repair and regeneration. Therefore, my working hypothesis in this proposal is that miR-302/367 functions as components of a Gata6-dependent regulatory network involving multiple positive and negative feedback loops to modulate proliferation and differentiation of lung epithelial progenitor cells. A better understanding of how miR-302/367 regulates the behaviors of lung epithelial progenitor cells during airway epithelium development and regeneration in response to injury will provide important insights into multiple lung diseases. This will be accomplished by pursuing two specific aims: Specific Aim 1. K99 Phase: Determine the role of miR-302/367 in modulating the behaviors of lung epithelial progenitor cells during airway epithelium development and regeneration after injury. a) Determine the effects of conditional gain- and loss-of-miR-302/367 on lung epithelial progenitor cell development using ROSA-miR- 302/367 and floxed miR-302/367 mouse lines. b) Determine whether decreased expression of Wnt5a is responsible for the expansion of lung epithelial progenitors in miR-302/367 gain of function mutants. c) Determine the requirement of miR-302/367 for homeostasis and regeneration in airway epithelium. Specific Aim 2. R00 Phase: Determine the mechanisms underlying the regulation of epithelial progenitor cell behaviors by miR-302/367 in airway epithelium development and regeneration. a) Characterize miR-302/367 expressing cells in lung epithelium during development and airway regeneration after injury. b) Determine the direct targets and downstream signaling pathways regulated by miR-302/367 in developing lung. c) Determine whether modulation of specific miR-302/367-regulated targets or pathways would enhance airway epithelium repair and regeneration.